The phenotype of vascular smooth muscle cells co-cultured with endothelial cells is modulated by PDGFR-β/IQGAP1 signaling in LPS-induced intravascular injury

Int. J. Med. Sci. 2019, Vol. 16

Ivyspring
International Publisher

1149

International Journal of Medical Sciences
2019; 16(8): 1149-1156. doi: 10.7150/ijms.34749

Research Paper

The phenotype of vascular smooth muscle cells
co-cultured with endothelial cells is modulated by
PDGFR-β/IQGAP1 signaling in LPS-induced
intravascular injury
Xia Zheng1* , Xiaotong Hu2*, Wang Zhang1
1.
2.

Department of Critical Care Medicine, The First Affiliated Hospital of Zhejiang University, Hangzhou, Zhejiang, 310003, P.R. China.
Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases; The First Affiliated Hospital, College of Medicine, Zhejiang University,
Hangzhou, Zhejiang, 310003, P.R. China.

*Co-first

author

 Corresponding author: X.Z. email: ; Postal address: 79 QingChun Road, Hangzhou, Zhejiang, 310003, P.R. China.
© The author(s). This is an open access article distributed under the terms of the Creative Commons Attribution License ( />See for full terms and conditions.

Received: 2019.03.10; Accepted: 2019.07.09; Published: 2019.08.06

Abstract
Background Sepsis, a leading cause of death in intensive care units, is generally associated with
vascular dysfunction. However, its pathophysiological process has not been fully clarified, lacking
in-depth knowledge of its pathophysiological process may hinder the improvement of diagnosis and
therapy for sepsis. Hence, as the key parts of the vascular wall, the interaction between endothelial
cells (ECs) and smooth muscle cells (SMCs) under septic situation need to be further studied.
Methods ECs and SMCs were co-cultured using Transwell plates. Lipopolysaccharide (LPS) was
used to induce sepsis. A scratch-wound assay was used to assess cell migration, and western blotting
was used to assess the level of redifferentiation of SMCs as well as the expression of PDGFR-β and
IQGAP1.
Results Co-culture with ECs reduced the redifferentiation of SMCs induced by LPS (10 μg/ml),
which was characterized by increased migration ability and decreased expression of contractile
proteins (e.g., SM22 and α-SMA). The production of TNF-α could decrease the level of PDGFR-β in
SMCs. Treatment of SMCs with the PDGFR-β inhibitor imatinib (5 μM) was able to counteract
LPS-induced SMC redifferentiation and reduce IQGAP1 protein expression, especially when SMCs
were co-cultured with ECs.
Conclusion The phenotype of vascular SMCs co-cultured with ECs was modulated by IQGAP1
through the PDGFR-β pathway, which may lead to vascular remodeling and homeostasis in
LPS-induced intravascular injury. This pathway could be a novel target for the treatment of vascular
damage.
Key words: sepsis; endothelial cells; smooth muscle cells; phenotype; Intravascular Injury.

Introduction
Sepsis is a major challenge in public health care.
Considerable resources have been invested in
developing potential therapies [1]. The current
treatment for sepsis is primarily symptomatic support
[2]. This is likely a result of the lack of in-depth

knowledge of the underlying pathophysiological

processes of sepsis. It is well known that vascular
dysfunction is a decisive factor in the development of
several inflammatory diseases. The mechanisms
underlying septic induction of oxidative and
nitrosative stresses, the functional consequences of
these stresses, and potential adjunct therapies for



Int. J. Med. Sci. 2019, Vol. 16
microvascular dysfunction have been identified [3].
As demonstrated in the literature, the interaction
between endothelial cells (ECs) and smooth muscle
cells (SMCs) is an essential maturation process in
physiological conditions [4, 5]. Generally, in vitro
studies are based on single cell cultures, which
exclude interactions between different cell types.
Previous studies have suggested that ECs could
regulate vascular tone through the release of
vasoactive molecules such as platelet-derived growth
factor (PDGF)-BB [6, 7] and tumor necrosis factor-α
(TNF-α) [8]. There are two types of SMCs: contractile
and synthetic SMCs. Synthetic SMCs have stronger
migratory ability compared with contractile SMCs [9].
Contractile and synthetic SMCs can be distinguished
by differences in the expression levels of marked
proteins, such as smooth muscle 22 (SM22) and
α-smooth muscle actin (α-SMA), which are known as

contractile SMC proteins. Studies have shown that the
PDGF receptor (PDGFR)-β pathway plays a key role
in SMC phenotypic modulation by suppressing the
expression of SM22 and α-SMA [10, 11], resulting in a
synthetic phenotype that can facilitate the infiltration
of inflammatory cells [12].
IQ-domain
GTPase-activating
protein
1
(IQGAP1) plays a key role in regulating cell migration
[13-15]. A previous study demonstrated that IQGAP1
expression was markedly increased in vascular
diseases caused by complete removal of the
endothelium [16], and that IQGAP1 played a critical
role in SMC migration at least in part by increasing
PDGFR in focal adhesions, as well as by increasing
focal adhesion formation at the leading edge [16].
However, the effects of IQGAP1 on SMC phenotypic
transformation and migration following vascular
damage caused by sepsis remain unknown. The
present study investigated the role of the
PDGFRβ/IQGAP1 pathway in EC-mediated SMC
phenotypic transformation and migration during
sepsis in a co-culture cell model.

Methods
Reagents
Lipopolysaccharide (LPS, Escherichia coli 055:
B5, Cat. No. L2880; Sigma-Aldrich) was used to mimic

a septic condition. LPS was diluted by
phosphate-buffered saline (PBS); The chemical
inhibitor imatinib mesylate (Cat. No. S1026; Selleck)
was used to inhibit the PDGFR; The first antibodies
included anti-IQGAP1 (1:1000; # ab86064, Abcam),
anti-α-SMA
(1:1000;
#A5228;
Sigma-Aldrich),
anti-SM22 (1:1000; #ab137453; Abcam), anti-GAPDH
(1:1000; #5174; Cell Signaling Technology); A
horseradish peroxidase (HRP)-conjugated secondary

1150
antibody was purchased
Technology (1:5000, #7074).

from

Cell

Signaling

Cell Culture and intervention
Human umbilical vein SMCs (Cat. No. 8020) and
ECs (Cat. No. 8000) were purchased from ScienCell.
SMCs were cultured in basal medium (SMCM, Cat.
No.1101; ScienCell), supplemented with 2% fetal
bovine serum (FBS, Cat. No. 0010; ScienCell), 1%
smooth muscle cell growth supplement (SCGS,

Cat.No.1152; ScienCell). ECs were cultured in basal
medium (ECM, Cat. No.1001; ScienCell) containing
5% fetal bovine serum (FBS, Cat. No. 0025; ScienCell),
and 1% endothelial cell growth supplement (ECGS,
Cat. No. 1052; ScienCell). After 1% penicillin
/streptomycin (P/S, Cat. No. 0503; ScienCell) was
added, they were maintained at 37 °C in a humidified
5% CO2 incubator. Passages 3–8 were used for the
experiments.

Co-culture of ECs and SMCs
The co-culture system was established by using
the Transwell plates (Cat. No. 3470; Corning)[17],
with SMCs were seeded in the lower wells and the
ECs were planted in the transwell inserts. Before ECs
were co-cultured with SMCs for 24 hours, they both
were separately pretreated as follows: culturing with
control vehicle (Control), culturing with LPS (LPS),
culturing with LPS and imatinib (LPS + imatinib)
according to different experiment design, imatinib
was given before LPS for 90 min, then SMCs and ECs
were extensively washed with PBS to remove excess
LPS and/or imatinib which were not taken up.
Serum-free mediums were added, in order to exclude
any confounding factors contained in the serum.

Enzyme-Linked immunosorbnent assay
Levels of PDGF-BB and TNF-α were determined
in the supernatants of different groups using
commercial high-sensitivity ELISAs, according to the

manufacturer’s instructions (Cat. No.EK91372,
MultiSciences Biotech, Co., Ltd).

Wound healing assay
SMCs were seeded in six-well plates according
to different groups. The cell monolayer was scratched
using a 200μl pipette tip before washing three times
with PBS to clear cell debris and floating cells. One
thousand microliters of serum-free SMCM was then
added, and the cells were incubated for 24 h at 37 °C
in 5% CO2. Images were captured under a microscope
before and after the 24 h incubation at the same
position. Migration ability was measured by
calculating the rate of scratch wound confluence after
24 h using Adobe Photoshop 2016 software (Adobe
Systems Inc.,).



Int. J. Med. Sci. 2019, Vol. 16
Western Blot Analysis
According to general procedure, western Blot
Analysis was performed, briefly, Equal amounts of
lysates of cells were applied to 4–12% SDS-PAGE
precast gels (Cat. No. NP0335, Invitrogen; Thermo
Fisher Scientific), Resolved proteins were transferred
to polyvinylidene fluoride (PVDF) membranes (Cat.
No. IPVH00010; Merck Millipore), blocked, and then
incubated with the primary and second antibody,
then the protein bands were visualized by enhanced

chemiluminescence
kit
(Cat.
No.
70-P1421;
MultiSciences Biotech, Co., Ltd.,) and exposed to
X-ray film. The expression of the protein was
analyzed by Image J software.

CCK-8 assay
Cells (1 × 105 cells/ml) were grown in 96-well
plates and then starved for 24 h before being subjected
to treatment according to the experimental
requirement, cells were then harvested and washed
with PBS and cell counting kit-8 (CCK-8; Dojindo)
mixed with FBS-free medium was used for cell
viability assay.

Statistical analysis
All results were shown as mean ± SD. Statistical
significance was assessed by unpaired Student’s t-test
or ANOVA, P-values of 0.01 and 0.05 were considered
significant. *p < 0.05, **p < 0.01.

Results
Co-culture of ECs and SMCs resulted in higher
SMC TNF-α expression
SMCs and ECs were treated according to the
experimental design shown in Figure 1. The
supernatants were collected from SMCs for an

1151
enzyme-linked immunosorbent assay (ELISA), which
indicated a slight upward trend in the level of TNF-α
in
single-culture
SMCs
treated
with
lipopolysaccharide (LPS); there was no significant
difference between the LPS-treated and control
groups. However, in the co-culture system, ECs
induced higher TNF-α expression in SMCs compared
with single-culture SMCs with or without LPS
treatment (mean ± standard deviation, 1077.37 ±
127.90 pg/ml vs. 187.47 ± 10.45 pg/ml; P < 0.01
without LPS treatment; 1907.69 ± 119.79 pg/ml vs.
284.17 ± 1.60 pg/ml; P < 0.01 with LPS treatment).
When the co-culture system was stimulated with LPS,
TNF-α expression reached the highest level of the four
subgroups (Figure 1A). There were no statistical
differences in the levels of PDGF-BB in the
above-mentioned groups (Figure 1B).

ECs affect the SMC phenotype in a paracrine
manner
To ascertain whether ECs can affect the SMC
phenotype in a paracrine manner, single-culture
SMCs were treated with control vehicle or LPS for 24
h. After the cell monolayer was scratched, serum-free

medium was added to the culture and the cells were
incubated for another 24 h. In the co-culture system,
SMCs and ECs were treated with or without LPS
separately for 24 h prior to co-culture, and the
subsequent steps were performed as described above
for the single-culture SMCs. The scratch-wound assay
revealed that LPS increased the migration ability of
SMCs compared with the control group in both the
single- and co-culture systems. When SMCs were
co-cultured with ECs, the increased migration
induced by LPS was alleviated. ECs had no influence
on the migration ability of SMCs in the absence of LPS
(Figure 2A and 2A1).

Figure 1. ECs induced higher TNF-α expression in SMCs. (A), The quantification of TNF-α expression in SMCs under single-culture or co-culture system. **: P<0.01,
SMCs were treated with control vehicle in single-culture system (SMC Control) vs SMCs were treated with control vehicle in co-culture system (CULSMC Control), ##: P<0.01,
SMCs were treated with LPS in single-culture (SMC LPS) vs SMCs were treated with LPS in co-culture (CULSMC LPS), @@: P<0.01, CULSMC Control vs CULSMC LPS; (B),
No significant difference in the level of PDGF-BB was shown in the above-mentioned groups.




Int. J. Med. Sci. 2019, Vol. 16

1152

Figure 2. SMCs Shift phenotypic transition to synthetic type from contractile type orchestrated by ECs. (A), LPS increased the migration ability of SMCs. When
SMCs were co-cultured with ECs, the increased migration induced by LPS was alleviated. (A1), Quantification of A is shown in A1, [% width of injury line = (a − b) × 100%/a; a
= Initial scratch wound area at 0 h, b = Scratch wound area at 24 h] **: P<0.01, SMC Control vs SMC LPS, ##: P<0.01, CULSMC Control vs CULSMC LPS, @@: P<0.01, SMC
LPS vs CULSMC LPS; (B), LPS induced the reduction of SM22 and α-SMA in SMCs, however, when SMCs were co-cultured with ECs, the reduction of SM22 and α-SMA protein

induced by LPS were partly alleviated. Meanwhile, ECs could enhance the expression of SM22 and α-SMA in SMCs in normal condition. (B1 and B2). Quantification of B is shown
in B1 and B2, **: P<0.01, SMC Control vs SMC LPS, ##: P<0.01, #: P<0.05, SMC LPS vs CULSMC LPS; CULSMC Control vs CULSMC LPS, @@: P<0.01, SMC LPS vs CULSMC
LPS; &: P<0.05, SMC Control vs CULSMC Control.

Western blotting demonstrated that LPS
treatment for 24 h eventually led to a phenotypic
transition characterized by the reduction of SM22 and
α-SMA in SMCs. When SMCs were co-cultured with
ECs, the protein levels of SM22 and α-SMA were
significantly increased; when treated with LPS, the
levels of SM22 and α-SMA were somewhat decreased.
Additionally, when exposed to LPS, SM22 and α-SMA
protein expression in SMCs co-cultured with ECs was
markedly higher compared with single-culture SMCs
(Figure 2B).
As shown in Figure 3A, when compared to
control groups, LPS induced a higher level of
phosphorylated PDGFR-β (p-PDGFR-β) in SMCs in
both the single and co-culture systems. ECs induced a

slight decrease in p-PDGFR-β levels of SMCs in the
absence of LPS treatment, but this difference was not
statistically significant. With LPS stimulation,
p-PDGFR-β levels in co-culture SMCs were
remarkably lower than those of single-culture SMCs.

IQGAP1 is involved in SMC differentiation in
the co-culture system
As illustrated by western blotting, increased
expression of IQGAP1 was observed in LPS-treated

SMCs in both the single and co-culture systems. In the
co-culture system, the increased expression of
IQGAP1 in LPS-treated SMCs was partly alleviated
(Figure 3B).




Int. J. Med. Sci. 2019, Vol. 16

1153

Figure 3. LPS induced a higher higher level of PDGFR-β and IQGAP1 in SMCs in both the single and co-culture systems. (A), LPS induced the level of
p-PDGFR-β expression in SMCs in both single and coculture system, but when SMCs were co-cultured with ECs, both control and LPS group experienced the relative lower
p-PDGFR-β protein expression than that in single culture system. With LPS stimulation, p-PDGFR-β levels in co-culture SMCs were remarkably lower than those of
single-culture SMCs. Quantification of A is shown in A1; **: P<0.01, SMC Control vs SMC LPS, #: P<0.05, CULSMC Control vs CULSMC LPS, @: P≤0.05, SMC LPS vs CULSMC
LPS; (B), the change of IQGAP1 expression was mediated by LPS, Quantification of B is shown in (B1); **: P<0.01, SMC Control vs SMC LPS, #: P<0.05, CULSMC Control vs
CULSMC LPS, @@: P<0.01, SMC LPS vs CULSMC LPS.

SMCs co-cultured in the absence of LPS were
treated with imatinib (Figure 3A). Imatinib had no
effect on the levels of p-PDGFR-β; however, as a
PDGFR antagonist, the effects of imatinib on LPS
induced migratory ability in SMCs. The phenotypic
transition of SMCs and expression of IQGAP1 in the
co-culture system were measured. We used imatinib
at 5 μM for the following experiments; this
concentration was confirmed by CCK8 assay (Figure
S1). In Figure 4, the scratch-wound assay and western
blotting showed that LPS induced a decrease in the

expression of α-SMA and SM22, and an increase in the
migration ability of SMCs in the co-culture system;
these effects were attenuated by the PDGFR-β
antagonist (Figure 4A and 4B). Next, IQGAP1 protein
expression was determined in the absence and
presence of imatinib. The results showed that LPS
induced increased expression of IQGAP1 in SMCs,
and imatinib prevented this increase in expression
(Figure 4C).

Discussion
The most characteristic aspects of sepsis may be
its complex pathophysiological processes, in which
vascular events are considered prominent; the role of
ECs in vascular events has been expanded immensely
[18-20]. However, SMCs are another component of
blood vessel with a less well-defined role during
sepsis, and the relationship between ECs and SMCs
has not yet been characterized.

A growing body of literature is indicating that
SMCs exhibit considerable phenotypic plasticity
[21-23]. When vascular damage occurs, constricted
SMCs are responsible for vascular contraction and
expansion, and can be transformed into synthetic
SMCs, which are characterized by decreased
expression of SM22 and α-SMA [21, 24, 25] and
increased migration ability. To date, few studies have
reported on the involvement of ECs in transformation
of the SMC phenotype during sepsis in a co-culture

system.
In the present study, LPS treatment resulted in a
decrease in the expression of contractile proteins and
an increased migration ability in single-culture SMCs,
and co-culture of ECs and SMCs resulted in
alleviation of the expression of contractile proteins
and suppression of migration ability in the presence
or absence of LPS. Together, our results provide
evidence that LPS induces SMC injury, and that ECs
could partly reverse the change in the phenotypic
transition and migration of SMCs.
PDGF is a potent mitogen for cells of
mesenchymal origin, including fibroblasts, smooth
muscle cells, and glial cells [26, 27]. In both mouse and
human, the PDGF signaling network consists of five
ligands, PDGF-AA through -DD (including -AB), and
two receptors, PDGFR-α and PDGFR-β. In general,
expression of PDGFRs is low in vivo, but increases
dramatically during inflammation. In human
pulmonary alveolar epithelial cells, LPS has been



Int. J. Med. Sci. 2019, Vol. 16
shown to induce p42/p44 MAPK activation via the
PDGFR/PI3K/Akt pathway [28]. Previous research
has revealed that PDGFR-β engages several
well-characterized signaling pathways known to be
involved in multiple LPS-induced cellular and
developmental responses [29-31]. Moreover, Oison et

al. found that PDGFR-β signaling is involved in the

1154
differentiation of vascular smooth muscle [21].
Similarly, in the present study, when compared to
control groups, LPS induced a higher level of
p-PDGFR-β in SMCs in both the single and co-culture
systems. In co-culture SMCs, imatinib had no effect on
p-PDGFR-β levels and could reverse the
LPS-mediated higher level of p-PDGFR-β in SMCs.

Figure 4. Imatinib attenuated vascular smooth muscle cell phenotype switching to a migration state by IQGAP1. (A), The changes of α-SMA and SM22 expression were
mediated by imatinib. Quantification of α-SMA is shown in A1; **: P<0.01, CULSMC Control vs CULSMC LPS, ##: P<0.01, CULSMC LPS vs SMCs were treated with LPS and
imatinib in co-culture system (CULSMC LPS + imatinib). Quantification of SM22 expression is shown in A2; *: P<0.05, CULSMC Control vs CULSMC LPS, #: P<0.05, CULSMC
LPS vs CULSMC LPS + imatinib; (B) LPS induced an increase in the migration ability of SMCs in the co-culture system were attenuated by imatinib. Quantification of B is shown
in B1, **: P<0.01, CULSMC Control vs CULSMC LPS, ##: P<0.01, CULSMC LPS vs CULSMC LPS + imatinib; (C) The change of IQGAP1 expression was mediated by imatinib.
Quantification of IQGAP1 is shown in C1; **: P<0.01, CULSMC Control vs CULSMC LPS, ##: P<0.01, CULSMC LPS vs CULSMC LPS + imatinib.




Int. J. Med. Sci. 2019, Vol. 16
A previous study found PDGFR-β could be
suppressed by TNF-α [32], which is often involved in
sepsis, and that TNF-α could reduce cell proliferation
in response to PDGF-BB [33]. In the present study,
TNF-α only exhibited a slight upward trend, while the
expression of PDGFR-β was significantly increased
following treatment with LPS, which may be affected
by multiple factors related to sepsis. In the absence of

LPS stimulation, ECs promoted an increase in TNF-α
expression in SMCs and tended to slightly decrease
PDGFR-β expression; with LPS stimulation, there was
an obvious increase in TNF-α expression and a
decrease
in
PDGFR-β
expression.
Thus,
TNF-α/PDGFR-β may be involved in the interaction
between ECs and SMCs in co-culture, especially with
LPS stimulation.
PDGF-BB is the highest affinity ligand for
PDGFR-β. In a recent study involving a co-culture
system, LPS-activated microglia stimulated PDGF-BB
expression, enhanced angiogenesis, migration,
proliferation, and permeability, and altered the
phenotype of co-cultured renal microvascular
endothelial cells [34]. Previous studies have
demonstrated that PDGF can be induced in ECs in
response to injury or stimulus and play a vital role in
SMCs and vessel remodeling [35, 36]. However, no
statistical differences were observed in PDGF-BB
levels between the different groups in this study. We
can speculate that not all PDGF family members were
detected, only PDGF-BB. A similar hypothesis was
proposed by Kim and colleagues; they determined
that PDGF-BB levels did not respond to LPS
treatment, but that PDGF-AA increased in a
dose-dependent manner with LPS stimulation [37].

Therefore, we suggest that the change in PDGFR-β
was affected by TNF-α rather than PDGF-BB in our
experiment.
In fact, a large corpus of data has shown that
PDGFR-β participates in SMC migration [38, 39].
Kohno and colleagues suggested that IQGAP1 might
contribute to SMC migration through interaction with
PDGFR-β [16]. IQGAP1 mediates protein–protein
interactions with a myriad of binding sites that
regulate numerous signaling pathways, contributing
to its multiple domains. For example, IQGAP1 forms
scaffolds with several components of the Akt [15] and
ERK [40, 41] pathways to facilitate diverse cellular
functions. Further evidence has suggested that Akt
signaling may regulate the SMC phenotype [23], and
that IQGAP1 operates at both the leading and trailing
edge of migrating cells via the ERK pathway [16].
These results prompted us to further explore the
regulation of IQGAP1 in response in injury and to
determine whether IQGAP1 was involved in SMC
differentiation. Consistent with the findings of

1155
previous studies [21, 42], increased expression of
IQGAP1 was observed in LPS-treated SMCs in both
the single and co-culture systems. Expression of
IQGAP1 was higher in single culture compared with
co-culture SMCs. In the co-culture system, the
increased expression of IQGAP1 in LPS-treated SMCs
was partly alleviated by the PDGFR-β inhibitor

imatinib. When SMCs were treated with imatinib, the
decreased expression of α-SMA and SM22, and
LPS-induced increased migration of co-cultured
SMCs, were attenuated. Thus, we conclude that LPS
could induce the phenotypic transition of SMCs to
migratory SMCs in the co-culture system via the
TNF-α/PDGFR-β/IQGAP1 pathway, and that the
different levels of these parameters in the single and
co-culture systems may be due to the interaction
between ECs and SMCs.

Conclusions
Our
findings
indicate
that
the
PDGFR-β/IQGAP1 pathway is involved in the
interaction between ECs and SMCs, and that TNF-α
may regulate PDGFR-β, especially in vascular injury
with LPS stimulation. Thus, the interaction between
ECs and SMCs serves an important role in vascular
homeostasis and remodeling during sepsis. Indeed,
this pathway may be a new target for the treatment of
vascular damage that occurs with sepsis. Further
study of additional signaling pathways involved in
EC-mediated SMC phenotypic transformation will be
necessary.

Supplementary Material

Supplementary figure S1.
/>
Acknowledgements
The work was supported by Young Science
Foundation of National Natural Science Foundation
of China (81101445) and Zhejiang Provincial Natural
Science Foundation (LY16H150002).

Competing Interests
The authors have declared that no competing
interest exists.

References
1.
2.
3.
4.
5.

Angus DC, Linde-Zwirble WT, Lidicker J, Clermont G, Carcillo J, Pinsky MR.
Epidemiology of severe sepsis in the United States: analysis of incidence,
outcome, and associated costs of care. Critical care medicine. 2001; 29: 1303-10.
Gotts JE, Matthay MA. Sepsis: pathophysiology and clinical management.
Bmj. 2016; 353: i1585.
Cepinskas G, Wilson JX. Inflammatory response in microvascular
endothelium in sepsis: role of oxidants. J Clin Biochem Nutr. 2008; 42: 175-84.
Jain RK. Molecular regulation of vessel maturation. Nature medicine. 2003; 9:
685-93.
Lao KH, Zeng L, Xu Q. Endothelial and smooth muscle cell transformation in
atherosclerosis. Curr Opin Lipidol. 2015; 26: 449-56.

Int. J. Med. Sci. 2019, Vol. 16
6.
7.
8.
9.
10.
11.

12.
13.
14.
15.

16.

17.
18.
19.
20.
21.
22.
23.
24.
25.
26.
27.

28.

29.

30.

31.
32.
33.
34.

Labrecque L, Lamy S, Chapus A, Mihoubi S, Durocher Y, Cass B, et al.
Combined inhibition of PDGF and VEGF receptors by ellagic acid, a
dietary-derived phenolic compound. Carcinogenesis. 2005; 26: 821-6.
Rolny C, Nilsson I, Magnusson P, Armulik A, Jakobsson L, Wentzel P, et al.
Platelet-derived growth factor receptor-beta promotes early endothelial cell
differentiation. Blood. 2006; 108: 1877-86.
Zhaowei J, D'Souza MJ, Oettinger CW. Reversal of LPS induced endothelial
cell TNF synthesis and increased permeability with microencapsulated
antisense oligomers to NF-kappaB. J Microencapsul. 2007; 24: 596-607.
Hao H, Gabbiani G, Bochaton-Piallat ML. Arterial smooth muscle cell
heterogeneity: implications for atherosclerosis and restenosis development.
Arteriosclerosis, thrombosis, and vascular biology. 2003; 23: 1510-20.
McDonald OG, Owens GK. Programming smooth muscle plasticity with
chromatin dynamics. Circ Res. 2007; 100: 1428-41.
Millette E, Rauch BH, Kenagy RD, Daum G, Clowes AW. Platelet-derived
growth factor-BB transactivates the fibroblast growth factor receptor to induce
proliferation in human smooth muscle cells. Trends Cardiovasc Med. 2006; 16:
25-8.
Rensen SS, Doevendans PA, van Eys GJ. Regulation and characteristics of

vascular smooth muscle cell phenotypic diversity. Neth Heart J. 2007; 15:
100-8.
Brown MD, Sacks DB. IQGAP1 in cellular signaling: bridging the GAP. Trends
Cell Biol. 2006; 16: 242-9.
Noritake J, Watanabe T, Sato K, Wang S, Kaibuchi K. IQGAP1: a key regulator
of adhesion and migration. Journal of cell science. 2005; 118: 2085-92.
Yamaoka-Tojo M, Ushio-Fukai M, Hilenski L, Dikalov SI, Chen YE, Tojo T, et
al. IQGAP1, a novel vascular endothelial growth factor receptor binding
protein, is involved in reactive oxygen species--dependent endothelial
migration and proliferation. Circ Res. 2004; 95: 276-83.
Kohno T, Urao N, Ashino T, Sudhahar V, Inomata H, Yamaoka-Tojo M, et al.
IQGAP1 links PDGF receptor-beta signal to focal adhesions involved in
vascular smooth muscle cell migration: role in neointimal formation after
vascular injury. American journal of physiology Cell physiology. 2013; 305:
C591-600.
Zhao L, Luo H, Li X, Li T, He J, Qi Q, et al. Exosomes Derived from Human
Pulmonary Artery Endothelial Cells Shift the Balance between Proliferation
and Apoptosis of Smooth Muscle Cells. Cardiology. 2017; 137: 43-53.
De Backer D, Ortiz JA, Salgado D. Coupling microcirculation to systemic
hemodynamics. Curr Opin Crit Care. 2010; 16: 250-4.
Koh IH, Menchaca-Diaz JL, Koh TH, Souza RL, Shu CM, Rogerio VE, et al.
Microcirculatory evaluation in sepsis: a difficult task. Shock. 2010; 34 Suppl 1:
27-33.
Levi M. Pathogenesis and treatment of disseminated intravascular coagulation
in the septic patient. Journal of critical care. 2001; 16: 167-77.
Olson LE, Soriano P. PDGFRbeta signaling regulates mural cell plasticity and
inhibits fat development. Dev Cell. 2011; 20: 815-26.
Cordes KR, Sheehy NT, White MP, Berry EC, Morton SU, Muth AN, et al.
miR-145 and miR-143 regulate smooth muscle cell fate and plasticity. Nature.
2009; 460: 705-10.

Tang R, Zhang G, Chen SY. Smooth Muscle Cell Proangiogenic Phenotype
Induced by Cyclopentenyl Cytosine Promotes Endothelial Cell Proliferation
and Migration. J Biol Chem. 2016; 291: 26913-21.
Sandbo N, Taurin S, Yau DM, Kregel S, Mitchell R, Dulin NO. Downregulation
of smooth muscle alpha-actin expression by bacterial lipopolysaccharide.
Cardiovasc Res. 2007; 74: 262-9.
Leimgruber C, Quintar AA, Sosa LD, Garcia LN, Figueredo M, Maldonado
CA. Dedifferentiation of prostate smooth muscle cells in response to bacterial
LPS. Prostate. 2011; 71: 1097-107.
Hannink M, Donoghue DJ. Structure and function of platelet-derived growth
factor (PDGF) and related proteins. Biochim Biophys Acta. 1989; 989: 1-10.
Heldin CH. Structural and functional studies on platelet-derived growth
factor. The EMBO journal. 1992; 11: 4251-9.
Cho RL, Yang CC, Lee IT, Lin CC, Chi PL, Hsiao LD, et al. Lipopolysaccharide
induces ICAM-1 expression via a c-Src/NADPH oxidase/ROS-dependent
NF-kappaB pathway in human pulmonary alveolar epithelial cells. Am J
Physiol Lung Cell Mol Physiol. 2016; 310: L639-57.
Ha T, Hua F, Liu X, Ma J, McMullen JR, Shioi T, et al.
Lipopolysaccharide-induced
myocardial
protection
against
ischaemia/reperfusion injury is mediated through a PI3K/Akt-dependent
mechanism. Cardiovasc Res. 2008; 78: 546-53.
Zhang L, Li HY, Li H, Zhao J, Su L, Zhang Y, et al. Lipopolysaccharide
activated phosphatidylcholine-specific phospholipase C and induced IL-8 and
MCP-1 production in vascular endothelial cells. J Cell Physiol. 2011; 226:
1694-701.
Severgnini M, Takahashi S, Rozo LM, Homer RJ, Kuhn C, Jhung JW, et al.
Activation of the STAT pathway in acute lung injury. Am J Physiol Lung Cell

Mol Physiol. 2004; 286: L1282-92.
Molander C, Kallin A, Izumi H, Ronnstrand L, Funa K. TNF-alpha suppresses
the PDGF beta-receptor kinase. Exp Cell Res. 2000; 258: 65-71.
Battegay EJ, Raines EW, Colbert T, Ross R. TNF-alpha stimulation of fibroblast
proliferation. Dependence on platelet-derived growth factor (PDGF) secretion
and alteration of PDGF receptor expression. J Immunol. 1995; 154: 6040-7.
Ding X, Gu R, Zhang M, Ren H, Shu Q, Xu G, et al. Microglia enhanced the
angiogenesis, migration and proliferation of co-cultured RMECs. BMC
Ophthalmol. 2018; 18: 249.

1156
35. Drusbosky L, Gars E, Trujillo A, McGee C, Meacham A, Wise E, et al.
Endothelial cell derived angiocrine support of acute myeloid leukemia
targeted by receptor tyrosine kinase inhibition. Leuk Res. 2015; 39: 984-9.
36. Lin CM, Chiu JH, Wu IH, Wang BW, Pan CM, Chen YH. Ferulic acid
augments angiogenesis via VEGF, PDGF and HIF-1 alpha. J Nutr Biochem.
2010; 21: 627-33.
37. Kim TH, Moon JH, Savard CE, Kuver R, Lee SP. Effects of lipopolysaccharide
on platelet-derived growth factor isoform and receptor expression in cultured
rat common bile duct fibroblasts and cholangiocytes. J Gastroenterol Hepatol.
2009; 24: 1218-25.
38. Tan M, Yan HB, Li JN, Li WK, Fu YY, Chen W, et al. Thrombin Stimulated
Platelet-Derived Exosomes Inhibit Platelet-Derived Growth Factor
Receptor-Beta Expression in Vascular Smooth Muscle Cells. Cell Physiol
Biochem. 2016; 38: 2348-65.
39. Cai Y, Nagel DJ, Zhou Q, Cygnar KD, Zhao H, Li F, et al. Role of
cAMP-phosphodiesterase 1C signaling in regulating growth factor receptor
stability, vascular smooth muscle cell growth, migration, and neointimal
hyperplasia. Circ Res. 2015; 116: 1120-32.
40. Bardwell AJ, Lagunes L, Zebarjedi R, Bardwell L. The WW domain of the

scaffolding protein IQGAP1 is neither necessary nor sufficient for binding to
the MAPKs ERK1 and ERK2. J Biol Chem. 2017; 292: 8750-61.
41. Sbroggio M, Bertero A, Velasco S, Fusella F, De Blasio E, Bahou WF, et al.
ERK1/2 activation in heart is controlled by melusin, focal adhesion kinase and
the scaffold protein IQGAP1. Journal of cell science. 2011; 124: 3515-24.
42. Yang P, Wu J, Miao L, Manaenko A, Matei N, Zhang Y, et al. Platelet-Derived
Growth Factor Receptor-beta Regulates Vascular Smooth Muscle Cell
Phenotypic Transformation and Neuroinflammation After Intracerebral
Hemorrhage in Mice. Critical care medicine. 2016; 44: e390-402.